Methods in Molecular Biology

0373375 - MBÚ 2012 RIV US eng M - Monography Chapter
Mašek, T. - Valášek, Leoš - Pospíšek, M.
Polysome Analysis and RNA Purification from Sucrose Gradients.
Methods in Molecular Biology. Vol. 703. New York: Springer Science + Business Media (Humana Press), 2011 - (Nielsen, H.), s. 293-309. ISBN 978-1-58829-913-0
R&D Projects: GA MŠMT LC06066
Institutional research plan: CEZ:AV0Z50200510
Keywords : Translational control * Polysome profile * RNA isolation
Subject RIV: CE - Biochemistry

Velocity separation of translation complexes in linear sucrose gradients is the ultimate method for both analysis of the overall fitness of protein synthesis as well as for detailed investigation of physiological roles played by individual factors of the translational machinery. Polysome profile analysis is a frequently performed task in translational control research that not only enables direct monitoring of the efficiency of translation but can easily be extended with a wide range of downstream applications such as Northern and Western blotting, genome-wide microarray analysis or qRT-PCR. This chapter provides a basic overview of the polysome profile analysis technique and the RNA isolation procedure from sucrose gradients. We also discuss possible experimental pitfalls of data normalization, describe main alternatives of the basic protocol and outline a novel application of denaturing RNA electrophoresis in several steps of polysome profile analysis
Permanent Link: http://hdl.handle.net/11104/0206485